cAMP sensor Epac as a determinant of ATP-sensitive potassium channel activity in human pancreatic β cells and rat INS-1 cells

The Epac family of cAMP-regulated guanine nucleotide exchange factors (cAMPGEFs, also known as Epac1 and Epac2) mediate stimulatory actions of the second messenger cAMP on insulin secretion from pancreatic β cells. Because Epac2 is reported to interact in vitro with the isolated nucleotide-binding fold-1 (NBF-1) of the β-cell sulphonylurea receptor-1 (SUR1), we hypothesized that cAMP might act via Epac1 and/or Epac2 to inhibit β-cell ATP-sensitive K + channels (K ATP channels; a hetero-octomer of SUR1 and Kir6.2). If so, Epac-mediated inhibition of K ATP channels might explain prior reports that cAMP-elevating agents promote β-cell depolarization, Ca 2 + influx and insulin secretion. Here we report that Epac-selective cAMP analogues (2′- O -Me-cAMP; 8-pCPT-2′- O -Me-cAMP; 8-pMeOPT-2′- O -Me-cAMP), but not a cGMP analogue (2′- O -Me-cGMP), inhibit the function of K ATP channels in human β cells and rat INS-1 insulin-secreting cells. Inhibition of K ATP channels is also observed when cAMP, itself, is administered intracellularly, whereas no such effect is observed upon administration N 6 -Bnz-cAMP, a cAMP analogue that activates protein kinase A (PKA) but not Epac. The inhibitory actions of Epac-selective cAMP analogues at K ATP channels are mimicked by a cAMP agonist (8-Bromoadenosine-3′, 5′-cyclic monophosphorothioate, Sp-isomer, Sp-8-Br-cAMPS), but not a cAMP antagonist (8-Bromoadenosine-3′, 5′-cyclic monophosphorothioate, Rp-isomer, Rp-8-Br-cAMPS), and are abrogated following transfection of INS-1 cells with a dominant-negative Epac1 that fails to bind cAMP. Because both Epac1 and Epac2 coimmunoprecipitate with full-length SUR1 in HEK cell lysates, such findings delineate a novel mechanism of second messenger signal transduction in which cAMP acts via Epac to modulate ion channel function, an effect measurable as the inhibition of K ATP channel activity in pancreatic β cells.